The present proposal is a competitive revision application for an R01 grant EY06177 entitled "Mechanism of Lacrimal Gland Secretion". The overall purpose of our research on lacrimal gland secretion has been to determine the signaling pathways that nerves use to stimulate lacrimal gland secretion under normal conditions, discover which steps in the signaling pathways are dysfunctional in dry eye disease, and develop treatments to reverse or repair the dysfunction. In the pursuit of these goals, we have identified a subset of cells in the lacrimal gland that can be cultured which have characteristics of stem or progenitor cells. The present proposal extends these findings and explores the possibility of isolating and transplanting these cells into diseased lacrimal glands. We plan to isolate and characterize lacrimal gland progenitor cells from adult tissue using GFP+ mice. Confocal microscopy will be performed with markers to stem cells and flow cytometry will be used to isolate these cells. Lacrimal glands from GFP+ mice will be grown in culture to determine if lacrimal gland progenitor cells form floating spheres (lacrispheres), which cell types are present in the lacrispheres, and which factors induce development of progenitor cells into functional, terminally differentiated lacrimal gland cell acinar, duct, myoepithelial, and nerve cells. These exogenous progenitor cells will then be injected into lacrimal glands from diseased mice to determine if they will repopulate the gland with functioning acinar, duct, myoepithelial, and nerve cells and restore the gland's secretory function. Finally, we will determine if allogeneic donor progenitor cells and/or their terminally differentiated offspring are targets of immune rejection in either diseased or healthy recipient lacrimal glands by determining 1) if mature terminally differentiated lacrimal gland cells derived from progenitor cells are immune privileged or immunogenic, 2) what immune effector cells are responsible for rejection or what immunosuppressive factors are responsible for immune privilege and 3) what immunosuppressive agents can prolong survival of either progenitor cells, and/or mature differentiated cells. We believe the use of lacrimal gland progenitor cells will allow us to develop treatments or potentially a cure for dry eye disease. PUBLIC HEALTH RELEVANCE: Millions of Americans suffer from dry eye disease from a variety of causes including aging, refractive surgery, menopause, autoimmune disease, and trauma causing damage to the ocular surface resulting in pain, inflammation, and irritation. There is no cure for dry eye and as the lacrimal gland is the primary contributor to the aqueous layer of the tear film, replacement of a diseased lacrimal gland using stem cells would improve the health of the ocular surface.